Genomic analysis is revolutionizing early disease diagnosis and dramatically enhancing patient care (McGuire et al. Science 317:1687, Srinivas et al., Lancet Oncol. 2:698). Microarrays (Drmanac et al., Science 260:1649, Hacia et al., Nat. Genet. 14:441) and polymerase chain reaction (PCR)-based techniques (Saiki et al., Science 230:1350) have, as tools, helped to spearhead this revolution, enabling the discovery and the initial development of assays for patient testing (Morris et al., Curr. Opin. Oncol. 19:547). However, spreading the reach of the genomics revolution to the patient bedside demands cost effective tools for individual biomarker profiling assessed relative to a posited disease state. Specifically, tools enabling routine patient care preferably would be simpler, more portable, and less expensive than PCR-based methods, yet should retain a high degree of selectivity and sensitivity.
Biomarker analysis based on electronic readout has long been cited as a promising approach that would enable a new family of chip-based devices with appropriate cost and sensitivity for medical testing (Drummond et al., Nat. Biotechnol. 21:1192, Katz et al., Electroanalysis 15: 913). The sensitivity of electronic readout is in principle sufficient to allow direct detection of small numbers of analyte molecules with simple instrumentation. However, despite tremendous advances in this area as well as related fields working towards new diagnostics (Clack et al., Nat. Biotechnol. 26:825, Geiss et al., Nat. Biotechnol. 2:317, Hahm et al., Nano Lett. 4:51, Munge et al., Anal. Chem. 77:4662, Nicewamer-Pena et al., Science 294:137, Park et al., Science 295:1503, Sinensky et al., Nat. Nano. 2:653, Steemers et al., Nat. Biotechnol. 18:91, Xiao et al., J. Am. Chem. Soc. 129:11896, Zhang et al., Nat. Nano. 1:214, Zhang et al., Anal. Chem. 76:4093, Yi et al., Biosens. Bioelectron. 20:1320, Ke et al., Science 319:180, Armani et al., Science 317:783), current multiplexed chips have yet to achieve direct electronic detection of biomarkers in cellular and clinical samples. The challenges that have limited the implementation of such devices primarily stem from the difficulty of obtaining very low detection limits in the presence of high background noise levels present when complex biological samples are assayed, and the challenge of generating multiplexed systems that are highly sensitive and specific.
The miniaturization of electrochemical systems continues to be a major focus in analytical and bioanalytical chemistry (Matysik, Miniaturization of Electroanalytical Systems (Springer-Verlag, 2002)), as the attainment of enhanced sensitivity may be enabled with systems possessing micro- to nano-scale dimensions (Szamocki et al., A. Anal. Chem. 2007, 79, 533-539). A great deal of work has been carried out with electrodes with dimensions on the micrometer or sub-micrometer scale. These systems offer many advantages over conventional macroelectrodes such as faster double-layer charging, reduced ohmic loss, high mass-transport rates, and high current density (Bond et al. Anal. Chimi. Acta 1989m 216, 177-230, Heinze, Angew. Chem. Int. Ed. 2003, 32, 1268-1288). Indeed, such electrodes have become well-established tools in a wide range of analytical applications (Bard, Electrochemical Methods: Fundamentals and Applications (Wiley, New York, 2001), Reimers, Chem. Rev. 2007, 107, 590-600, Zosic, Handbook of electrochemistry (Elsevier, 2007)). Working with nanoscale electrodes, however, is significantly more challenging, as fabrication is typically labour-intensive, insufficiently reproducible, and the currents obtained from such structures are typically difficult to measure accurately.
The use of nanowire electrodes for ultrasensitive nucleic acids and protein detection has been investigated (Gasparac et al. J Am Chem Soc 726:12270). The use of this electrode platform enables the electrochemical detection of picomolar levels of analytes, a level of sensitivity that is not possible using macroscale materials. Although it has been reported that nanowires are able to detect attomolar levels of analytes, this actually corresponds to picomolar levels when dealing with the volumes typically used for analysis. It has also been demonstrated that nanoparticle-modified electrodes may exhibit several advantages over conventional macroelectrodes such as enhancement of mass transport, catalysis, high effective surface area and control over electrode microenvironment (Katz et al. Electroanalysis 2004, 16, 19-44, Welch et al. Anal. Bioanal. Chem. 2006, 384, 601-619). Manufacturing arrays of nanowire electrodes, however, is non-trivial.
Boron doped diamond microelectrodes modified by electrodeposition of platinum nanoparticles have been used for the oxidative determination of As(III) at levels below 1 ppb (Hrapovic et al. Anal. Chem. 2007, 79, 500-507). However, this type of electrode cannot be incorporated into an array-based format for multiplexed experiments.
The analysis of panels of nucleic acid or protein biomarkers offers valuable diagnostic and prognostic information for clinical decision making. Existing methods that offer the specificity and sensitivity to profile clinical samples are typically costly, slow and serial. There is thus a need for an ultrasensitive device for detecting biomarkers in a multiplexed fashion.